Monitoring Binding of HIV-1 Capsid Assembly Inhibitors Using 19F Ligand-and 15N Protein-Based NMR and X-ray Crystallography: Early Hit Validation of a Benzodiazepine Series

作者：Goudreau Nathalie^*; Coulombe Rene; Faucher Anne Marie; Grand Maitre Chantal; Lacoste Jean Eric; Lemke Christopher T; Malenfant Eric; Bousquet Yves; Fader Lee; Simoneau Bruno; Mercier Jean Francois; Titolo Steve; Mason Stephen W

来源：ChemMedChem, 2013, 8(3): 405-414.

DOI：10.1002/cmdc.201200580

• binding studies
• fluorine
• HIV-1 capsids
• NMR spectroscopy
• X-ray crystallography

The emergence of resistance to existing classes of antiretroviral drugs underlines the need to find novel human immunodeficiency virus (HIV)-1 targets for drug discovery. The viral capsid protein (CA) represents one such potential target. Recently, a series of benzodiazepine inhibitors was identified via high-throughput screening using an invitro capsid assembly assay (CAA). Here, we demonstrate how a combination of NMR and X-ray co-crystallography allowed for the rapid characterization of the early hits from this inhibitor series. Ligand-based 19FNMR was used to confirm inhibitor binding specificity and reversibility as well as to identify the N-terminal domain of the capsid (CANTD) as its molecular target. Protein-based NMR (1H and 15N chemical shift perturbation analysis) identified key residues within the CANTD involved in inhibitor binding, while X-ray co-crystallography confirmed the inhibitor binding site and its binding mode. Based on these results, two conformationally restricted cyclic inhibitors were designed to further validate the possible binding modes. These studies were crucial to early hit confirmation and subsequent lead optimization.